Lung transplants have been performed in over 20,000 individuals with advanced lung disease, with two-year survival of 70% and dramatic improvements in quality of life. However, long-term survival rates remain lower than those of kidney, heart, and liver recipients, largely due to the effects of acute and chronic lung rejection (bronchiolitis obliterans syndrome, BOS). Current methods of predicting and diagnosing lung rejection are imprecise and invasive, and do not allow diagnosis of chronic rejection prior to irreversible loss of lung function. The lung allograft is ideally suited to study by bronchoalveolar lavage (BAL), which allows relatively non-invasive sampling of the bronchoalveolar microenvironment. Therefore, our objective is to use gene expression in BAL cells to improve prediction, diagnosis, and treatment of lung rejection. Our prior studies demonstrate that acute lung rejection histology is characterized by coordinately increased BAL cell expression of cytotoxic T-lymphocyte markers;and that BAL fluid and cells reveal biomarkers of the hallmark patholophysiologic processes in chronic lung rejection. To expand upon these findings we propose two hypotheses, each with a related Specific Aim. Hypothesis 1: Specific patterns of bronchoalveolar lavage (BAL) cell gene expression occur during acute rejection, and therefore can serve as diagnostic biomarkers. In Specific Aim 1, we will refine and validate our acute lung rejection diagnostic model by: a) adjusting for the effects on gene expression of common bronchial colonizing organisms;and b) validating the final acute lung rejection diagnostic model in a longitudinal, prospective study of lung recipients. Successful completion of Specific Aim 1 will pave the way to use BAL as the primary acute rejection diagnostic tool, reserving more invasive lung biopsies for problematic cases. Hypothesis 2: Specific biological mechanisms involved in BOS pathogenesis--including activation of innate and adaptive immunity;activation of neutrophil/chemokine-mediated inflammatory pathways;abnormal epithelial cell signaling;and airway fibrosis-are reflected in BAL cell gene expression. In Specific Aim 2, we will define the BAL cell expression of gene modules that reflect specific biological mechanisms involved in bronchiolitis obliterans syndrome pathogenesis by: a) evaluating the expression of known mechanism-based gene modules at the time of BOS onset;and b) identifying novel genes and pathways that can serve as candidate biomarkers for BOS. Successful completion of Specific Aim 2 will provide an empirical basis for development of mechanism-based diagnostic and predictive tests for BOS. These findings will be of immediate use, and will improve the clinical management of lung transplant recipients. We are uniquely positioned to carry out the proposed research based on our history of patient-oriented, proteomic, and genomic research in lung transplantation;our strong investigative team;and our outstanding genomic research environment. PUBLIC HEALTH RELEVANCE: Lung transplants have been performed in over 20,000 individuals with advanced lung disease, with two-year survival of approximately 70% and dramatic improvements in quality of life. However, long-term survival rates are considerably lower than those enjoyed by kidney, heart, and liver recipients, largely due to the effects of acute and chronic lung rejection. Our overall research goal is to reduce the negative impact of rejection after lung transplantation. In the proposed studies, we will study gene expression in lung cells in order to develop new ways of identifying lung rejection before it results in irreversible damage to the transplanted lungs.